Worldwide efforts to determine the genomic DNA sequences of humans and other animals are ongoing. Such efforts typically focus on obtaining sequence information from cDNAs in libraries created from RNAs of various tissues. Thus, collections of “expressed sequence tags” (ESTs) include portions of coding regions from most human genes.
Although ESTs provide useful structural information, they offer little insight into the functional relationship among genes. The functional relationship is of particular importance to determining the set of genes involved in a biological process and, subsequently, to developing pharmaceutical agents that affect one or more of the components of the biological process. See, e.g., Friedrich, G. A., “Moving Beyond the Genome Projects: Does the Future of Genomics-Based Drug Discovery Lie With the Mouse?,” Nature Biotechnology 14:1234-1237 (1996).
Friedrich argues in favor of using model systems that mirror human physiology in determining which genes may be involved in a biological process, and suggests that the mouse is an excellent model organism for human biology in that it shares with humans most salient aspects of mammalian physiology. The genomes of mice and humans are approximately the same in size, organization, and structure. Friedrich proposes that the mouse can be developed as an effective tool for drug development. Friedrich puts forth a “radical” suggestion that there is no logical barrier hindering large-scale phenotypic screens using mice.
Friedrich proposes using an insertional mutagen in embryonic stem cells to generate random mutations in the mouse genome, then screening for a variety of pre-determined phenotypes and cloning affected genes.
In particular, the physiology of, and treatments for, colon cancer are of particular biomedical interest. Colon cancer is one of the most prevalent malignancies in the Western world, with an estimated 145,000 new cases and 60,000 deaths each year in the United States alone. Genetic factors play a key role in this disease. Mutations in the human adenomatous polyposis coli (APC) gene cause a set of familial colon cancer syndromes. Mice carrying a mutation in a corresponding gene (Apc) also develop many intestinal adenomas. Heterozygotes for the Min (Multiple Intestinal Neoplasia) allele of the mouse Apc gene develop numerous intestinal and colonic adenomas [on average 29 Å 10, on a C57BL/6J (or equivalent derivative) background] that are similar in morphology to the adenomas seen in human inherited colonic polyposis syndromes such as familial adenomatous polyposis and Gardner's syndrome. Min/Min homozygotes die in utero. The Min mutation maps to mouse chromosome 18. The sequence of the Apc gene is known and published. Min mice carry a nonsense mutation in exon 15 of the mouse Apc gene (a mutation of the sort typically seen in human colon cancer kindreds). Mice carrying Min thus provide a model system for studying human familial adenomatous polyposis.
A locus (Mom-1) that strongly modifies the tumor number in heterozygous Min/+ mice was mapped to distal chromosome 4. Dietrich, W. F., et al., “Genetic Identification of Mom-1, a major modifier locus affecting Min-induced intestinal neoplasia in the mouse,” Cell 75:631-639 (1993). Mom-1 lies in a region of synteny conservation with human chromosome 1p35-36, a region of frequent somatic loss of heterozygosity in a variety of human tumors, including colon tumors. Mom-1 is only one of an unknown number of Loci that modify the expression of an inherited cancer syndrome, and it does not explain all of the genetic variation in tumor number in intraspecific backcrosses.
What is lacking is a systematic method for pinpointing genetic loci involved in modifying known phenotypes, by enhancing or suppressing. In the particular case of colon cancer in humans and animals, it would be desirable to locate the sequences in the genome (and the molecules encoded by those sequences) that are involved in the appearance of intestinal adenomas. The lack of such a systematic method has limited understanding of oncogenesis and, as such, has precluded development of pharmaceuticals that modify the oncogenic process. A systematic method should include not only non-essential loci, for which numerous mutant alleles can be found among homozygous inbred mouse strains, but also essential loci, for which mutant alleles in heterozygous form may influence the phenotype. Mutations that inactivate an essential gene will normally be lethal when homozygous, and so will not be found among inbred mouse strains.